Production and characterization of two N-terminal truncated esterases from Thermus thermophilus HB27 in a mesophilic yeast: Effect of N-terminus in thermal activity and stability

Fuciños, Pablo; Atanes, Estrella; López-López, Olalla; Cerdán, M. Esperanza; González-Siso, María-Isabel; Pastrana, Lorenzo; Rúa, M. Luisa

doi:10.1016/j.pep.2011.04.002

Cited by 18 publications

(21 citation statements)

References 33 publications

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“…3A, the esterase had maximum activity at pH 7.5. Similar results were reported earlier for esterases from Brevibacterium linens ATCC 9174 [24] and Thermus thermophilus HB27 [25]. Effect of temperature on the enzyme activity was determined at different temperatures ranging from 20 to 80°C.…”

Section: Effect Of Ph and Temperature On Enzyme Activitysupporting

confidence: 80%

Purification and characterization of a pH and heat stable esterase from Geobacillus sp. TF17

Çolak

2013

Turk J Bioch

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Section: Effect Of Ph and Temperature On Enzyme Activitysupporting

confidence: 80%

Purification and characterization of a pH and heat stable esterase from Geobacillus sp. TF17

Çolak

2013

Turk J Bioch

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“…Although these enzymes are active at high temperatures, they appear to be less stable over time when incubated at elevated temperatures. Furthermore, thermostable lipases have been reported in different Thermus isolates [19], [20] and were recently expressed in thermophilic yeasts [21], [22]. A thermostable esterase from Thermus scotoductus has been reported that was partially biochemically characterized [23].…”

Section: Introductionmentioning

confidence: 99%

The Metagenome-Derived Enzymes LipS and LipT Increase the Diversity of Known Lipases

et al. 2012

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Triacylglycerol lipases (EC 3.1.1.3) catalyze both hydrolysis and synthesis reactions with a broad spectrum of substrates rendering them especially suitable for many biotechnological applications. Most lipases used today originate from mesophilic organisms and are susceptible to thermal denaturation whereas only few possess high thermotolerance. Here, we report on the identification and characterization of two novel thermostable bacterial lipases identified by functional metagenomic screenings. Metagenomic libraries were constructed from enrichment cultures maintained at 65 to 75°C and screened resulting in the identification of initially 10 clones with lipolytic activities. Subsequently, two ORFs were identified encoding lipases, LipS and LipT. Comparative sequence analyses suggested that both enzymes are members of novel lipase families. LipS is a 30.2 kDa protein and revealed a half-life of 48 h at 70°C. The lipT gene encoded for a multimeric enzyme with a half-life of 3 h at 70°C. LipS had an optimum temperature at 70°C and LipT at 75°C. Both enzymes catalyzed hydrolysis of long-chain (C12 and C14) fatty acid esters and additionally hydrolyzed a number of industry-relevant substrates. LipS was highly specific for (R)-ibuprofen-phenyl ester with an enantiomeric excess (ee) of 99%. Furthermore, LipS was able to synthesize 1-propyl laurate and 1-tetradecyl myristate at 70°C with rates similar to those of the lipase CalB from Candida antarctica. LipS represents the first example of a thermostable metagenome-derived lipase with significant synthesis activities. Its X-ray structure was solved with a resolution of 1.99 Å revealing an unusually compact lid structure.

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“…Pf_Est presented optimum pH activity at 6.5-7.0 in naphthol-derivate substrates, which is slightly lower than the optimum pH (7.5) reported earlier for esterases from Brevibacterium linens ATCC 9174, Thermus thermophilus HB27, and Geobacillus sp. TF17 [34][35][36]. The optimum temperature for Pf_Est activity was 80°C in a-naphthyl acetate (and butyrate) and 75°C in pNPP.…”

Section: Discussionmentioning

confidence: 99%

Characterization and Low-Resolution Structure of an Extremely Thermostable Esterase of Potential Biotechnological Interest from Pyrococcus furiosus

et al. 2016

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Enzymes isolated from extremophiles often exhibit superior performance and potential industrial applications. There are several advantages performing biocatalysis at elevated temperatures, including enhanced reaction rates, increased substrate solubility and decreased risks of contamination. Furthermore, thermophilic enzymes usually exhibit high resistance against many organic solvents and detergents, and are also more resistant to proteolytic attack. In this study, we subcloned and characterized an esterase from the hyperthermophilic archaeon Pyrococcus furiosus (Pf_Est) that exhibits optimal activity around 80 °C using naphthol-derived substrates and p-nitrophenyl palmitate (pNPP). According to the circular dichroism spectra, the secondary structure of P. furiosus esterase, which is predominantly formed by a β-sheet structure, is very stable, even after incubation at 120°C. We performed SAXS to determine the low-resolution structure of Pf_Est, which is monomeric in solution at 80 °C and has a molecular weight of 28 kDa. The Km and V values for this esterase acting on pNPP were 0.53 mmol/L and 6.5 × 10 U, respectively. Pf_Est was most active in the immiscible solvents and retained more than 50 % in miscible solvents. Moreover, Pf_Est possesses transesterification capacity, presenting better results when isobutanol was used as an acyl acceptor (2.69 ± 0.14 × 10 μmol/min mg) and the highest hydrolytic activity toward olive oil among different types of oils testes in this study. Collectively, these biophysical and catalytic properties are of interest for several biotechnological applications that require harsh conditions, including high temperature and the presence of organic solvents.

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Production and characterization of two N-terminal truncated esterases from Thermus thermophilus HB27 in a mesophilic yeast: Effect of N-terminus in thermal activity and stability

Cited by 18 publications

References 33 publications

Purification and characterization of a pH and heat stable esterase from Geobacillus sp. TF17

Purification and characterization of a pH and heat stable esterase from Geobacillus sp. TF17

The Metagenome-Derived Enzymes LipS and LipT Increase the Diversity of Known Lipases

Characterization and Low-Resolution Structure of an Extremely Thermostable Esterase of Potential Biotechnological Interest from Pyrococcus furiosus

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